Apple Calli Research Articles

Organization and regulation of the apple SUMOylation system under salt and ABA

Small ubiquitin-related modifier (SUMO)-mediated post-translational protein modification is widely conserved among eukaryotes. SUMOylation refers to the covalent attachment of SUMO to target proteins that alters their function, location, and protein–protein interactions when plants are under abiotic stress. We identified 37 genes in the apple genome that encoded members of the SUMOylation pathway. In addition, RNA-Seq data shows their expression levels between different tissues. We can find that there are mainly expressed genes between each component to ensure that the entire pathway works in the plant. We found that the expression levels of 12 genes were significantly changed under NaCl and ABA treatment through qRT-PCR. MdSIZ1a strongly expression responded to NaCl and ABA treatment. Subsequently, MdSIZ1a was cloned and transformed into apple callus, further verifying the important role of the SUMOylation pathway under stress conditions. The interaction between MdSIZ1a and MdSCEa was verified by yeast two-hybrid, confirming that MdSIZ1a acts as bridge enzyme on MdSCEa and target substrates. Finally, we predicted and analyzed the functional interaction network of E3 ligase to shed light on protein interactions and gene regulatory networks associated with DNA damage repair under abiotic stress in apples.

Transcriptome Analysis of Apples in High-Temperature Treatments Reveals a Role of MdLBD37 in the Inhibition of Anthocyanin Accumulation.

Coloring in apple fruit due to anthocyanin accumulation is inhibited by high temperature; however, the underlying mechanism remains unclear. In the present study, total anthocyanin and cyanidin 3-galactoside contents were determined and compared between cv. ‘Redchief Delicious’ apple fruits at 25 °C and 35 °C treatments. The high temperature (35 °C) treatment substantially decreased total anthocyanin and cyanidin 3-galactoside contents. The transcriptomes of 25 °C- and 35 °C-treated apples were analyzed by high-throughput RNA sequencing. A total of 8354 differentially expressed genes (DEGs) were detected at four time points corresponding to the two temperature treatments. The up-regulated DEGs were annotated using GO as well as KEGG databases. A network module of 528 genes (including 21 transcription factors) most associated with the total anthocyanin and cyanidin 3-galactoside contents was constructed by weighted correlation network analysis (WGCNA). In the WGCNA module, we unearthed a LOB domain-containing gene designated as MdLBD37. The expression of MdLBD37 was sharply up-regulated by high temperature and negatively correlated with the total anthocyanin and cyanidin 3-galactoside contents. Overexpression of MdLBD37 in apple fruit and calli decreased the expression of anthocyanin biosynthetic genes, such as MdCHI, MdCHS, MdF3H, MdANS, MdDFR, and MdUFGT, along with anthocyanin accumulation. Our results suggested that MdLBD37 significantly influenced the high-temperature inhibition of anthocyanin accumulation in apples. The findings shed more light on the mechanism of anthocyanin inhibition during high-temperature stress in apples.

Genome-wide identification and characterization of the Prunus persica ferredoxin gene family and its role in improving heat tolerance

Ferredoxin is involved in many biological processes, such as carbon fixation, nitrogen assimilation, chlorophyll metabolism, and fatty acid synthesis, and it plays a role in plant resistance to stress. However, the functions of Fds in peach during stress are unclear. In this study, 11 members of the peach Fd gene family were identified and divided into six groups (I– VI). We carried out bioinformatics analysis on these sequences, analyzed the physical and chemical properties of PpFd protein and the cis-elements in its promoter region, and predicted and compared the differences in gene structure and conserved protein motifs among groups. The results showed that the PpFd protein was highly conserved in plant species. In addition, overexpression of PpFd08 significantly increased the tolerance of transgenic tomato to high-temperature stress. The transcriptome analysis and qRT-PCR results of PpFd08 transgenic apple calli showed that PpFd08 might enhance heat resistance by modulating the expression of heat tolerance related genes. The results of this study provide a new understanding for the further study of the function of PpFd protein in peach and a candidate gene for improving the heat resistance of peach.

Genome-wide Identification, Expression, and Functional Analysis of MdMSI Genes in Apples (Malus domestica Borkh.).

The multicopy suppressor of IRA (MSI) is a subfamily of WD40 repeat proteins, which is widely involved in plant growth and development. In order to explore the function of MdMSI members in abiotic stress, we identified eight MSI gene family members from the Malus × domestica reference genome. They were distributed on six chromosomes, and they had similar secondary and tertiary structures. We found a variety of regulatory elements in response to hormones and abiotic stress in MdMSI promoters. Through qRT-PCR analysis, it was revealed that MdMSIs were expressed in all tissues, especially in roots. The analysis results also revealed that the expression of MdMSIs was induced in varying degrees under salt, drought stress, and ABA treatments. Furthermore, we obtained the overexpression of MdMSI1-1 transgenic apple calli and Arabidopsis. The overexpression of MdMSI1-1 in calli and Arabidopsis played a negative regulatory role in salt stress response. Our work laid a foundation for further verifying the function of MSI genes under abiotic stress in apples.

BTB protein MdBT2 negatively regulates iron homeostasis by interacting with MdNAC1 in apple

NAC transcription factors play an important role in plant abiotic stress response. However, the molecular mechanism of how it regulates iron accumulation remain largely unknown in apple. Here, we show that MdNAC1 positively regulates iron accumulation under iron deficiency conditions. Overexpression of MdNAC1 show higher levels of iron accumulation and chlorophyll content as well as activated the activity of plasma membrane (PM) H + -ATPases in response to iron deficiency. Quantitative PCR (qPCR) analysis showed that the expression of several iron-related genes were significantly up-regulated in transgenic apple calli. Additionally, we identify an MdBT2 protein that interacts with the MdNAC1 protein. MdBT2 negatively regulates the protein stability of MdNAC1 by degrading it through the ubiquitinated 26S proteasome pathway. In addition, overexpression of MdBT2 in apple and Arabidopsis exhibited inhibition of iron accumulation and showed chlorosis in the new leaves. In sum, our study provided molecular and genetic evidence that the MdBT2 may negatively regulate iron response by mediating the MdNAC1 stability. • We identified an NAC transcription factor, MdNAC1, from apple and demonstrated that it was localized in the nucleus. • The over expression of MdNAC1 increased iron accumulation in transgenic apple calli and Arabidopsis. • MdBT2 interacts with MdNAC1and mediates the ubiquitination of MdNAC1.

PpNUDX8, a Peach NUDIX Hydrolase, Plays a Negative Regulator in Response to Drought Stress.

Drought stress is a serious abiotic stress source that affects the growth and fruit quality of peach trees. However, the molecular mechanism of the NUDIX hydrolase family in peaches in response to drought stress is still unclear. Here, we isolated and identified the PpNUDX8 (Prupe.5G062300.1) gene from the peach NUDIX hydrolase family, and found that PpNUDX8 has a typical NUDIX hydrolase domain. In this study, we performed 15% PEG6000 drought treatment on peach seedlings, and qRT–PCR analysis showed that 15% PEG6000 induced the transcription level of PpNUDX8. Overexpression of PpNUDX8 reduced the tolerance of calli to 4% PEG6000 treatment. Compared with wild-type apple calli, PpNUDX8 transgenic apple calli had a lower fresh weight and higher MDA content. After 15% PEG6000 drought treatment, PpNUDX8 transgenic tobacco had a greater degree of wilting and shorter primary roots than Under control conditions. The chlorophyll, soluble protein, and proline contents in the transgenic tobacco decreased, and the MDA content and relative conductivity increased. At the same time, PpNUDX8 negatively regulated ABA signal transduction and reduced the transcriptional expression of stress response genes. In addition, PpNUDX8 was not sensitive to ABA, overexpression of PpNUDX8 reduced the expression of the ABA synthesis-related gene NCED6 and increases the expression of the ABA decomposition-related gene CYP1 in tobacco, which in turn leads to a decrease in the ABA content in tobacco. In addition, Under control conditions, overexpression of PpNUDX8 destroyed the homeostasis of NAD and reduced nicotinamide adenine dinucleotide (NADH) in tobacco. After 15% PEG6000 drought treatment, the changes in NAD and NADH in PpNUDX8 transgenic tobacco were more severe than those in WT tobacco. In addition, PpNUDX8 also interacted with PpSnRk1γ (Prupe.6G323700.1).

Overexpression of MdZAT5, an C2H2-Type Zinc Finger Protein, Regulates Anthocyanin Accumulation and Salt Stress Response in Apple Calli and Arabidopsis.

Zinc finger proteins are widely involved and play an important role in plant growth and abiotic stress. In this research, MdZAT5, a gene encoding C2H2-type zinc finger protein, was cloned and investigated. The MdZAT5 was highly expressed in flower tissues by qRT-PCR analyses and GUS staining. Promoter analysis showed that MdZAT5 contained multiple response elements, and the expression levels of MdZAT5 were induced by various abiotic stress treatments. Overexpression of MdZAT5 in apple calli positively regulated anthocyanin accumulation by activating the expressions of anthocyanin biosynthesis-related genes. Overexpression of MdZAT5 in Arabidopsis also enhanced the accumulation of anthocyanin. In addition, MdZAT5 increased the sensitivity to salt stress in apple calli. Ectopic expression of MdZAT5 in Arabidopsis reduced the expression of salt-stress-related genes (AtNHX1 and AtABI1) and improved the sensitivity to salt stress. In conclusion, these results suggest that MdZAT5 plays a positive regulatory role in anthocyanin accumulation and negatively regulates salt resistance.

Transcription factor McWRKY71 induced by ozone stress regulates anthocyanin and proanthocyanidin biosynthesis in Malus crabapple

In plants, anthocyanins and proanthocyanidins (PAs) play important roles in plant resistance to abiotic stress. In this study, ozone (O3) treatments caused the up-regulation of Malus crabapple structural genes McANS, McCHI, McANR and McF3H, which promoted anthocyanin and PA accumulation. We identified the WRKY transcription factor (TF) McWRKY71 by screening differentially expressed genes (DEGs) that were highly expressed in response to O3 stress from an RNA sequencing (RNA-seq) analysis. Overexpressing McWRKY71 increased the resistance of ‘Orin’ apple calli to O3 stress and promoted the accumulation of anthocyanins and PAs, which facilitated reactive oxygen species scavenging to further enhance O3 tolerance. Biochemical and molecular analyses showed that McWRKY71 interacted with McMYB12 and directly bound the McANR promoter to participate in the regulation of PA biosynthesis. These findings provide new insights into the WRKY TFs mechanisms that regulate the biosynthesis of secondary metabolites, which respond to O3 stress, in Malus crabapple.

MdWRKY126 modulates malate accumulation in apple fruit by regulating cytosolic malate dehydrogenase (MdMDH5).

The content of organic acids greatly influences the taste and storage life of fleshy fruit. Our current understanding of the molecular mechanism of organic acid accumulation in apple (Malus domestica) fruit focuses on the aluminum-activated malate transporter 9/Ma1 gene. In this study, we identified a candidate gene, MdWRKY126, for controlling fruit acidity independent of Ma1 using homozygous recessive mutants of Ma1, namely Belle de Boskoop "BSKP" and Aifeng "AF." Analyses of transgenic apple calli and flesh and tomato (Solanum lycopersicum) fruit demonstrated that MdWRKY126 was substantially associated with malate content. MdWRKY126 was directly bound to the promoter of the cytoplasmic NAD-dependent malate dehydrogenase MdMDH5 and promoted its expression, thereby enhancing the malate content of apple fruit. In MdWRKY126 overexpressing calli, the mRNA levels of malate-associated transporters and proton pump genes also significantly increased, which contributed to the transport of malate accumulated in the cytoplasm to the vacuole. These findings demonstrated that MdWRKY126 regulates malate anabolism in the cytoplasm and coordinates the transport between cytoplasm and vacuole to regulate malate accumulation. Our study provides useful information to improve our understanding of the complex mechanism regulating apple fruit acidity.

A MdMa13 gene encoding tonoplast P3B-type ATPase regulates organic acid accumulation in apple

The organic acid content is one of the important factors that determine fruit quality. In this study, one gene encoding a P3B-ATPase (designated as MdMa13) was identified as a determinant of fruit acidity in apple. Subcellular localization indicated that MdMa13 was located in the tonoplast. Overexpression of MdMa13 in apple calli, tomato or apples led to increased malic acid contents. In the transgenic apple calli, the expression levels of seven acid-related genes were higher than that in the wild-type apple calli, including MdMa1, MdtDT, MdDTC1, MdDTC2, MdVHA-A3, MdVHA-D, and MdMa11. The expression level of MdMa10 was lower in transgenic lines than that in wild-type apple calli. Together, these findings implied that MdMa13 has important functions related to apple fruit acidity. The results of this study will strengthen the characterization for the complex molecular regulation of fruit acidity.

Apple receptor-like kinase FERONIA regulates salt tolerance and ABA sensitivity in Malus domestica

FERONIA (FER) is a membrane-localized receptor-like kinase that plays pivotal roles in male and female gametophyte recognition, hormone signaling crosstalk, and biotic and abiotic responses. Most reports focus on the functions of FER in model plant Arabidopsis thaliana. However, the functions of FER homologs have not been deeply investigated in apple (Malus domestica), an important economic fruit crop distributed worldwide, especially in China. In this study, we identified an apple homolog of Arabidopsis FER, named MdFER (MDP0000390677). The two proteins encoded by AtFER and MdFER share similar domains: an extracellular malectin-like domain, a transmembrane domain, and an intracellular kinase domain. MdFER was further proven to localize to the plasma membrane in the epidermal cells of Nicotiana benthamiana. MdFER was widely expressed in different apple tissues, but the highest expression was found in roots. In addition, expression of MdFER was significantly induced by treatment with abscisic acid (ABA) and salt (NaCl). Overexpressing MdFER dramatically improved the resistance to salt stress and reduced the sensitivity to ABA in apple callus, while suppressing MdFER expression showed contrary effects. Furthermore, ectopic expression of MdFER in Arabidopsis significantly increased the salt tolerance and reduced the sensitivity to ABA. In addition, under salt stress and ABA treatment, Arabidopsis with highly expressed MdFER accumulated less reactive oxygen species (ROS), and the enzymatic activity of two ROS scavengers, superoxide dismutase and catalase, was higher compared with that of wild type (WT). Our work proves that MdFER positively regulates salt tolerance and negatively regulates ABA sensitivity in apple, which enriched the functions of FER in different plant species.

Overexpression of apple Ma12, a mitochondrial pyrophosphatase pump gene, leads to malic acid accumulation and the upregulation of malate dehydrogenase in tomato and apple calli.

Acidity is an important factor influencing the organoleptic quality of apple fruits. In this study, an apple pyrophosphate-energized proton pump (PEPP) gene was isolated and designated MdMa12. On the basis of a phylogenetic analysis in Rosaceae species, PEPP genes were divided into three groups, with apple PEPP genes most closely related to pear PEPP genes. Gene expression analysis revealed that high malic acid content was generally accompanied by high MdMa12 expression levels. Moreover, MdMa12 was mainly expressed in the fruit. A subcellular localization analysis suggested that MdMa12 is a mitochondrial protein. The ectopic expression and overexpression of MdMa12 in "Micro-Tom" tomato and apple calli, respectively, increased the malic acid content. One (MDH12) of four malate dehydrogenase genes highly expressed in transgenic apple calli was confirmed to encode a protein localized in mitochondria. The overexpression of MDH12 increased the malate content in apple calli. Furthermore, MdMa12 overexpression increased MdDTC1, MdMa1, and MdMa10 expression levels, which were identified to transport malate. These findings imply that MdMa12 has important functions related to apple fruit acidity. Our study explored the regulatory effects of mitochondria on the complex mechanism underlying apple fruit acidity.

Malate metabolism mediated by the cytoplasmic malate dehydrogenase gene MdcyMDH affects sucrose synthesis in apple fruit.

The types and proportions of soluble sugar and organic acid in fruit significantly affect flavor quality. However, there are few reports on the crosstalk regulation between metabolism of organic acid and sugar in fruit. Here, we found that the overexpression of cytoplasmic malate dehydrogenase genes (MdcyMDHs) not only increased the malate content but also increased the sucrose concentration in transgenic apple calli and mature fruit. Enzyme activity assays indicated that the overexpression of MdcyMDH1 and MdcyMDH5 enhanced sucrose phosphate synthase (SPS) activity in transgenic materials. RNA-seq and expression analysis showed that the expression levels of SPS genes were up-regulated in MdcyMDH1-overexpressed apple fruit and MdcyMDH5-overexpressed apple calli. Further study showed that the inhibition of MdSPSB2 or MdSPSC2 expression in MdcyMDH1 transgenic fruit could reduce or eliminate, respectively, the positive effect of MdcyMDH1 on sucrose accumulation. Moreover, some starch cleavage-related genes (MdBAM6.1/6.2, MdBMY8.1/8.2, MdISA1) and the key gluconeogenesis-related phosphoenolpyruvate carboxykinase MdPEPCK1 gene were significantly up-regulated in the transcriptome differentially expressed genes of mature fruit overexpressing MdcyMDH1. These results indicate that alteration of malate metabolism mediated by MdcyMDH might regulate the expression of MdSPSs and SPS activity via affecting starch metabolism or gluconeogenesis, and thus accelerate sucrose synthesis and accumulation in fruit.

Quantitative proteomic sequencing of F 1 hybrid populations reveals the function of sorbitol in apple resistance to Botryosphaeria dothidea.

Apple ring rot, which is caused by Botryosphaeria dothidea, is one of the most devastating diseases of apple. However, the lack of a known molecular resistance mechanism limits the development of resistance breeding. Here, the ‘Golden Delicious’ and ‘Fuji Nagafu No. 2’ apple cultivars were crossed, and a population of 194 F1 individuals was generated. The hybrids were divided into five categories according to their differences in B. dothidea resistance during three consecutive years. Quantitative proteomic sequencing was performed to analyze the molecular mechanism of the apple response to B. dothidea infection. Hierarchical clustering and weighted gene coexpression network analysis revealed that photosynthesis was significantly correlated with the resistance of apple to B. dothidea. The level of chlorophyll fluorescence in apple functional leaves increased progressively as the level of disease resistance improved. However, the content of soluble sugar decreased with the improvement of disease resistance. Further research revealed that sorbitol, the primary photosynthetic product, played major roles in apple resistance to B. dothidea. Increasing the content of sorbitol by overexpressing MdS6PDH1 dramatically enhanced resistance of apple calli to B. dothidea by activating the expression of salicylic acid signaling pathway-related genes. However, decreasing the content of sorbitol by silencing MdS6PDH1 showed the opposite phenotype. Furthermore, exogenous sorbitol treatment partially restored the resistance of MdS6PDH1-RNAi lines to B. dothidea. Taken together, these findings reveal that sorbitol is an important metabolite that regulates the resistance of apple to B. dothidea and offer new insights into the mechanism of plant resistance to pathogens.

Identification of MdGRF genes and the necessary role of MdGRF02 in apple root growth regulation

Growth-regulating factors (GRFs) as plant-specific transcription factors exert an important role in plant growth and development; however, the function of GRFs has not been reported in apple (Malus domestica). In the present study, we identified 12 MdGRF genes, which located on six chromosomes with different densities. These MdGRFs were phylogenetically classified into four clusters based on the protein sequence similarity, and the intron-exon structure and functional domain of MdGRF genes were similar within each cluster. Interestingly, these MdGRFs were tissue-specific expressed, and several specific MdGRFs were strongly responsive to various hormonal treatments, either positively or negatively. The transgenic apple calli overexpressing MdGRF02 displayed higher fresh weight and the transgenic apple roots exhibited increased root elongation, indicating that MdGRF02 may be related with root growth regulation in apples. This study reports the identification and expression analysis of the MdGRFs and provides some useful information to reveal the potential roles of MdGRFs in apple growth regulation, especially the root growth.

Glucose sensor MdHXK1 activates an immune response to the fungal pathogen Botryosphaeria dothidea in apple.

Sugars are essential regulatory molecules involved in plant growth and development and defense response. Although the relationship between sugars and disease resistance has been widely discussed, the underlying molecular mechanisms remain unexplored. Ring rot caused by Botryosphaeria dothidea (B. dothidea), which severely affects fruit quality and yield, is a destructive disease of apples (Malus domestica Borkh.). The present study found that the degree of disease resistance in apple fruit was closely related to glucose content. Therefore, the gene encoding a hexokinase, MdHXK1, was isolated from the apple cultivar 'Gala', and characterized during the defense response. Overexpression of MdHXK1 enhanced disease resistance in apple calli, leaves and fruits by increasing the expression levels of genes related to salicylate (SA) synthesis (PHYTOALEXIN DEFICIENT 4, PAD4; PHENYLALANINE AMMONIA-LYASE, PAL; and ENHANCED DISEASE SUSCEPTIBILITY 1, EDS1) and signaling (PR1; PR5; and NONEXPRESSER OF PR GENES 1, NPR1) as well as increasing the superoxide (O2- ) production rate and the hydrogen peroxide (H2 O2 ) content. Overall, the study provides new insights into the MdHXK1-mediated molecular mechanisms by which glucose signaling regulates apple ring rot resistance.

Expression Analysis of AUX/IAA Family Genes in Apple Under Salt Stress.

Members of the auxin/indoleacetic acid (Aux/IAA) gene family in plants are primary auxin-responsive genes that play important roles in many aspects of plant development and in responses to abiotic stress. Recently, 33 Aux/IAA have been identified in the apple genome. The biological responses of MdIAAs to salt stress are still unknown. In this study, Malus zumi, Malus baccata, and Malus × domestica 'Fuji' plantlets were subjected to salt stress by supplementing hydroponic media with NaCl at various concentrations. M. zumi showed the strongest salt resistance, followed by 'Fuji', and M. baccata was the most sensitive to salt stress. Tissue-specific expression profiles of MdIAAs were determined by quantitative real-time polymerase chain reaction. When apple plantlets were subjected to salt stress, most of salt-responsive MdIAAs were up-regulated by 1h, 3h, and 6h in roots, shoot tips, and leaves, respectively. Highly expressed MdIAAs in roots, especially for M. zumi, consisted with the salt tolerance of apple rootstocks. Transgenic apple calli were tolerant to salt stress when over-expressed salt-responsive genes, MdIAA8, -9, and -25. These results provide clues about salt resistance in these three Malus species, which helps apple breeding of salt tolerance by genetic transformation.

Genome-Wide Analysis of the Apple CBL Family Reveals That Mdcbl10.1 Functions Positively in Modulating Apple Salt Tolerance.

Abiotic stresses are increasingly harmful to crop yield and quality. Calcium and its signaling pathway play an important role in modulating plant stress tolerance. As specific Ca2+ sensors, calcineurin B-like (CBL) proteins play vital roles in plant stress response and calcium signaling. The CBL family has been identified in many plant species; however, the characterization of the CBL family and the functional study of apple MdCBL proteins in salt response have yet to be conducted in apple. In this study, 11 MdCBL genes were identified from the apple genome. The coding sequences of these MdCBL genes were cloned, and the gene structure and conserved motifs were analyzed in detail. The phylogenetic analysis indicated that these MdCBL proteins could be divided into four groups. The functional identification in Na+-sensitive yeast mutant showed that the overexpression of seven MdCBL genes could confer enhanced salt stress resistance in transgenic yeast. The function of MdCBL10.1 in regulating salt tolerance was also verified in cisgenic apple calli and apple plants. These results provided valuable insights for future research examining the function and mechanism of CBL proteins in regulating apple salt tolerance.

The PyPIF5-PymiR156a-PySPL9-PyMYB114/MYB10 module regulates light-induced anthocyanin biosynthesis in red pear

Some cultivars of pear (Pyrus L.) show attractive red fruit skin due to anthocyanin accumulation. This pigmentation can be affected by environmental conditions, especially light. To explore the light-induced regulation network for anthocyanin biosynthesis and fruit coloration in pear, small RNA libraries and mRNA libraries from fruit skins of ‘Yunhongyihao’ pear were constructed to compare the difference between bagging and debagging treatments. Analysis of RNA-seq of fruit skins with limited light (bagged) and exposed to light (debagged), showed that PyPIF5 was down-regulated after bag removal. PymiR156a was also differentially expressed between bagged and debagged fruit skins. We found that PyPIF5 negatively regulated PymiR156a expression in bagged fruits by directly binding to the G-box motif in its promoter. In addition, PymiR156a overexpression promoted anthocyanin accumulation in both pear skin and apple calli. We confirmed that PymiR156a mediated the cleavage of PySPL9, and that the target PySPL9 protein could form heterodimers with two key anthocyanin regulators (PyMYB114/PyMYB10). We proposed a new module of PyPIF5-PymiR156a-PySPL9-PyMYB114/MYB10. When the bagged fruits were re-exposed to light, PyPIF5 was down-regulated and its inhibitory effect on PymiR156a was weakened, which leads to degradation of the target PySPL, thus eliminating the blocking effect of PySPL on the formation of the regulatory MYB complexes. Ultimately, this promotes anthocyanin biosynthesis in pear skin.

Integrated Analysis of the Metabolome and Transcriptome on Anthocyanin Biosynthesis in Four Developmental Stages of Cerasus humilis Peel Coloration.

Cerasus humilis is a unique dwarf shrub and fruit color is an important trait in the species. In this study, we evaluated the transcriptomic and metabolomic profiles of the plant at different developmental stages to elucidate the mechanism underlying color formation. In a metabolomics analysis, 16 anthocyanin components were identified at four developmental stages, and high levels of cyanidin O-syringic acid and pelargonidin 3-O-beta-d-glucoside (callitephin chloride) were correlated with the reddening of the fruit peel. A co-expression analysis revealed that ANS and UFGT play key roles in pigmentation (PCC > 0.82). Additionally, transcriptome data showed that most anthocyanin biosynthetic genes and two MYB transcription factors were significantly up-regulated. QRT-PCR results for these differentially expressed genes were generally consistent with the high-throughput sequencing. Moreover, the overexpression of ChMYB1 (TRINITY_DN21536_c0_g1) in apple calli could contribute to the accumulation of anthocyanin. It was also found that UFGT (TRINITY_DN19893_c1_g5) and ChMYB1 (TRINITY_DN21536_c0_g1) have similar expression patterns. These findings provide insight into the mechanisms underlying anthocyanin accumulation and coloration during fruit peel development, providing a basis for the breeding of anthocyanin-rich C. humilis cultivars.